Abstract Project 4 will test the hypothesis that computationally designed protein ?minibinders? and logic-gated switches targeting cerebrospinal fluid (CSF) protein biomarkers and protein particles can serve as versatile capture and detection agents to vastly improve the molecular characterization of CSF samples. The availability of these reagents should support the overall U19 goal of improving our understanding of CSF biomarkers as direct measures of age-related cognitive decline and Alzheimer's Disease (AD) pathophysiology. We have integrated our work plan within the highly focused U19 Project: Next Generation Translational Proteomics for Alzheimer's and Related Dementias to test the above hypothesis. In collaboration with Projects 1?3, our main goal is to develop an optimized set of computationally designed minibinders (hyperstable binding proteins of length <65 aa) as CSF protein biomarker capture agents, and ultra-specific logic-gated reagents for detection of CSF particles that have two defined protein components. The research will iterate between computational design and experimental testing, with feedback at each stage from CSF assay experiments conducted in collaboration with Projects 1-3 which will guide improvement of the minibinder design methods for capture of specified CSF biomarker protein targets, and for ultra-specific logic- gated detection of CSF particles with two composite protein components. The outcomes will be (i) specific CSF biomarker capture and detection systems for AD and other age-related neurodegenerative disorders, and (ii) an integrated computational-experimental pipeline for rapid on demand engineering of new protein based diagnostic agents for neurodegenerative disorders in general. Therefore, Project 4 relies on a close collaboration with Projects 1-3 and Cores 1-4 within the highly interactive U19 program.